Biological manganese removal process has a problem of long start-up period, when it is adopted to purify groundwater containing high concentration of iron, manganese and ammonia nitrogen. In order to investigate the effect of circulation reflux on start-up, three starting modes with varying reflux ratio, constant reflux ratio and no reflux were adopted in three same filters. Results showed that the total iron, manganese and ammomia nitrogen in effluent of the three filters was below 0.3 mg/L, 0.05 mg/L and 0.2 mg/L on 1,1 and 82 day, respectively; Variation of iron, manganese and ammonia nitrogen removal along the filter showed that the iron was removed mainly in the depth of 0-0.4 m, the manganese was adsorbed by manganese sand at the beginning and biological manganese removal was improved gradually after ammonia nitrogen reduced to be low enough. The circulation reflux is an effective way to shorten the start-up period of biological manganese removal process with high concentration of iron, manganese and ammonia nitrogen.